BACKGROUND: Many factors are associated with the development and progression of liver fat and fibrosis; however, genetics and the gut microbiota are representative factors. Moreover, recent studies have indicated a link between host genes and the gut microbiota. This study investigated the effect of patatin-like phospholipase domain-containing 3 (PNPLA3) rs738409 (C > G), which has been reported to be most involved in the onset and progression of fatty liver, on liver fat and fibrosis in a cohort study related to gut microbiota in a non-fatty liver population. METHODS: This cohort study included 214 participants from the health check-up project in 2018 and 2022 who had non-fatty liver with controlled attenuation parameter (CAP) values <248 dB/m by FibroScan and were non-drinkers. Changes in CAP values and liver stiffness measurement (LSM), liver-related items, and gut microbiota from 2018 to 2022 were investigated separately for PNPLA3 rs738409 CC, CG, and GG genotypes. RESULTS: Baseline values showed no difference among the PNPLA3 rs738409 genotypes for any of the measurement items. From 2018 to 2022, the PNPLA3 rs738409 CG and GG genotype groups showed a significant increase in CAP and body mass index; no significant change was observed in the CC genotype group. LSM increased in all genotypes, but the rate of increase was highest in the GG genotype, followed by the CG and CC genotypes. Fasting blood glucose levels increased in all genotypes; however, HOMA-IR (Homeostasis Model Assessment of Insulin Resistance) increased significantly only in the GG genotype. HDL (high-density lipoprotein) and LDL (low-density lipoprotein) cholesterol levels significantly increased in all genotypes, whereas triglycerides did not show any significant changes in any genotype. As for the gut microbiota, the relative abundance of Feacalibacterium in the PNPLA3 rs738409 GG genotype decreased by 2% over 4 years, more than 2-fold compared to CC and GG genotypes. Blautia increased significantly in the CC group. CONCLUSION The results suggest that PNPLA3 G-allele carriers of non-fatty liver develop liver fat and fibrosis due to not only obesity and insulin resistance but also the deterioration of gut microbiota, which may require a relatively long course of time, even years.
Humans ; Gastrointestinal Microbiome ; Male ; Female ; Membrane Proteins/metabolism* ; Lipase/genetics* ; Middle Aged ; Liver Cirrhosis/epidemiology* ; Cohort Studies ; Genotype ; Adult ; Non-alcoholic Fatty Liver Disease/microbiology* ; Polymorphism, Single Nucleotide ; Acyltransferases ; Phospholipases A2, Calcium-Independent

OBJECTIVE: To investigate the therapeutic potential of pseudolaric acid B (PAB) on non-alcoholic fatty liver disease (NAFLD) and its underlying molecular mechanism in vitro and in vivo. METHODS: Eight-week-old male C57BL/6J mice (n=32) were fed either a normal chow diet (NCD) or a high-fat diet (HFD) for 8 weeks. The HFD mice were divided into 3 groups according to a simple random method, including HFD, PAB low-dose [10 mg/(kg·d), PAB-L], and PAB high-dose [20 mg/(kg·d), PAB-H] groups. After 8 weeks of treatment, glucose metabolism and insulin resistance were assessed by oral glucose tolerance test (OGTT) and insulin tolerance test (ITT). Biochemical assays were used to measure the serum and cellular levels of total cholesterol (TC), triglycerides (TG), aspartate aminotransferase (AST), alanine aminotransferase (ALT), low-density lipoprotein cholesterol (LDL-C), and high-density lipoprotein cholesterol (HDL-C). White adipose tissue (WAT), brown adipose tissue (BAT) and liver tissue were subjected to hematoxylin and eosin (H&E) staining or Oil Red O staining to observe the alterations in adipose tissue and liver injury. PharmMapper and DisGeNet were used to predict the NAFLD-related PAB targets. Peroxisome proliferator-activated receptor alpha (PPARα) pathway involvement was suggested by Kyoto Encyclopedia of Genes and Genomes (KEGG) and search tool Retrieval of Interacting Genes (STRING) analyses. Luciferase reporter assay, cellular thermal shift assay (CETSA), and drug affinity responsive target stability assay (DARTS) were conducted to confirm direct binding of PAB with PPARα. Molecular dynamics simulations were applied to further validate target engagement. RT-qPCR and Western blot were performed to assess the downstream genes and proteins expression, and validated by PPARα inhibitor MK886. RESULTS: PAB significantly reduced serum TC, TG, LDL-C, AST, and ALT levels, and increased HDL-C level in HFD mice (P<0.01). Target prediction analysis indicated a significant correlation between PAB and PPARα pathway. PAB direct target binding with PPARα was confirmed through luciferase reporter assay, CETSA, and DARTS (P<0.05 or P<0.01). The target engagement between PAB and PPARα protein was further confirmed by molecular dynamics simulations and the top 3 amino acid residues, LEU321, MET355, and PHE273 showed the most significant changes in mutational energy. Subsequently, PAB upregulated the genes expressions involved in lipid metabolism and mitochondrial biogenesis downstream of PPARα (P<0.05 or P<0.01). Significantly, the PPARα inhibitor MK886 effectively reversed the lipid-lowering and PPARα activation properties of PAB (P<0.05 or P<0.01). CONCLUSION PAB mitigates lipid accumulation, ameliorates liver damage, and improves mitochondrial biogenesis by binding with PPARα, thus presenting a potential candidate for pharmaceutical development in the treatment of NAFLD.
Animals ; PPAR alpha/metabolism* ; Non-alcoholic Fatty Liver Disease/pathology* ; Male ; Mice, Inbred C57BL ; Lipid Metabolism/drug effects* ; Diterpenes/therapeutic use* ; Organelle Biogenesis ; Diet, High-Fat ; Humans ; Mice ; Liver/metabolism* ; Insulin Resistance ; Mitochondria/metabolism* ; Molecular Docking Simulation

Non-alcoholic fatty liver disease (NAFLD), including non-alcoholic fatty liver (NAFL), non-alcoholic steatohepatitis (NASH), and advanced fibrosis, is a leading cause of chronic liver disease worldwide, progressing to cirrhosis and ultimately hepatocellular carcinoma (HCC). Excessive accumulation of fatty acids in the liver triggers multiple forms of hepatocyte death and exacerbates NAFLD progression, with pyroptosis and apoptosis considered key events. Recent studies show that cysteine aspartic acid specific protease-3 (caspase-3) is a central regulator of both pyroptosis and apoptosis in NAFLD. Activated caspase-3 not only directly induces apoptosis but also cleaves the N-terminal domain of gasdermin E (GSDME), disrupts cell membranes, releases inflammatory factors, and thereby mediates pyroptosis. Inhibiting caspase-3 expression in NAFLD can alleviate hepatocyte injury (such as ballooning degeneration), dampen pro-inflammatory signaling, and reduce apoptosis. Caspase-3 acts as a key node coordinating pyroptosis and apoptosis and may serve as a novel therapeutic target for the prevention and treatment of NAFLD.
Non-alcoholic Fatty Liver Disease/metabolism* ; Humans ; Pyroptosis/physiology* ; Apoptosis/physiology* ; Caspase 3/physiology* ; Animals ; Gasdermins

The gut microbiota is an indispensable symbiotic entity within the human holobiont, serving as a critical regulator of host lipid metabolism homeostasis. Therefore, it has emerged as a central subject of research in the pathophysiology of metabolic disorders. This microbial consortium orchestrates key aspects of host lipid dynamics-including absorption, metabolism, and storage-through multifaceted mechanisms such as the enzymatic processing of dietary polysaccharides, the facilitation of long-chain fatty acid uptake by intestinal epithelial cells (IECs), and the bidirectional modulation of adipose tissue functionality. Mounting evidence underscores that gut microbiota-derived metabolites not only directly mediate canonical lipid metabolic pathways but also interface with host immune pathways, epigenetic machinery, and circadian regulatory systems, thereby establishing an intricate crosstalk that coordinates systemic metabolic outputs. Perturbations in microbial composition (dysbiosis) drive pathological disruptions to lipid homeostasis, serving as a pathogenic driver for conditions such as obesity, hyperlipidemia, and non-alcoholic fatty liver disease (NAFLD). This review systematically examines the emerging mechanistic insights into the gut microbiota-mediated regulation of intestinal lipid metabolism, while it elucidates its translational implications for understanding metabolic disease pathogenesis and developing targeted therapies.
Humans ; Gastrointestinal Microbiome/physiology* ; Lipid Metabolism ; Animals ; Intestinal Mucosa/metabolism* ; Homeostasis ; Dysbiosis ; Obesity/metabolism* ; Intestines/microbiology* ; Non-alcoholic Fatty Liver Disease/metabolism* ; Metabolic Diseases/metabolism*

Nuclear factor erythroid 2-related factor 2 (Nrf2) is an intracellular transcription factor that helps protect against oxidative stress in different types of cells under pathological conditions. Mitochondria are vital organelles that function in diverse metabolic processes in the body, including redox reactions, lipid metabolism, and cell death. Mitophagy, a specific form of autophagy for damaged mitochondria, plays a critical role in the pathophysiology of liver diseases. In this review, we explain in detail the roles of the Nrf2 signaling pathway and mitophagy, and the relationship between them, in various hepatic diseases (nonalcoholic fatty liver disease, viral hepatitis, alcoholic liver disease, drug-induced liver injury, autoimmune hepatitis, hepatic ischemia‒reperfusion injury, and liver cancer). We also offer some potential insights and treatments relevant to clinical applications.
Humans ; NF-E2-Related Factor 2/metabolism* ; Mitophagy/physiology* ; Kelch-Like ECH-Associated Protein 1/metabolism* ; Signal Transduction ; Liver Diseases/etiology* ; Animals ; Oxidative Stress ; Mitochondria/metabolism* ; Non-alcoholic Fatty Liver Disease ; Liver Neoplasms

OBJECTIVES: To investigate the regulatory role of nucleotide-bound oligomerized domain-like receptor containing pyrin-domain protein 6 (NLRP6) in liver lipid metabolism and non-alcoholic fatty liver disease (NAFLD). METHODS: Mouse models with high-fat diet (HFD) feeding for 16 weeks (n=6) or with methionine choline-deficient diet (MCD) feeding for 8 weeks (n=6) were examined for the development of NAFLD using HE and oil red O staining, and hepatic expressions of NLRP6 were detected with RT-qPCR, Western blotting, and immunohistochemical staining. Cultured human hepatocytes (LO2 cells) with adenovirus-mediated NLRP6 overexpression or knock-down were treated with palmitic acid (PA) in the presence or absence of compound C (an AMPK inhibitor), and the changes in cellular lipid metabolism were examined by measuring triglyceride, ATP and β-hydroxybutyrate levels and using oil red staining, RT-qPCR, and Western blotting. RESULTS: HFD and MCD feeding both resulted in the development of NAFLD in mice, which showed significantly decreased NLRP6 expression in the liver. In PA-treated LO2 cells, NLRP6 overexpression significantly decreased cellular TG content and lipid deposition, while NLRP6 knockdown caused the opposite effects. NLRP6 overexpression in PA-treated LO2 cells also increased mRNA and protein expressions of PGC1A and CPT1A, levels of ATP and β-hydroxybutyrate, and the phosphorylation level of AMPK pathway; the oxidative decomposition of lipids induced by Ad-NLRP6 was inhibited by the use of AMPK inhibitors. CONCLUSIONS NLRP6 overexpression promotes lipid oxidation and decomposition through AMPK/CPT1A/PGC1A to alleviate lipid deposition in hepatocytes.
Non-alcoholic Fatty Liver Disease/metabolism* ; Animals ; Hepatocytes/metabolism* ; Lipid Metabolism ; Mice ; Humans ; Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha ; AMP-Activated Protein Kinases/metabolism* ; Carnitine O-Palmitoyltransferase/metabolism* ; Diet, High-Fat ; Male ; Mice, Inbred C57BL ; Signal Transduction

OBJECTIVES: To investigate the effect of Ching Shum Pills (CSP) for alleviating non-alcoholic fatty liver disease (NAFLD) and the underlying mechanism. METHODS: In a mouse model of NAFLD, the therapeutic effect of CSP was evaluated by measuring serum glucose, lipid profiles (TC, TG, LDL-C, HDL-C), and hepatic function markers. Network pharmacology was employed to identify active compounds in CSP and their targets using TCMSP, HERB, SwissTargetPrediction, GeneCards, OMIM, and DisGeNET. Protein-protein interaction (PPI) networks, Gene Ontology (GO), and KEGG pathway analyses were conducted. Molecular docking (AutoDock Vina) was used to assess the compound-target binding affinities. Quantitative real-time PCR (qRT-PCR) was used to validate the mRNA expressions of the core genes in the liver tissue of the mouse models. RESULTS: In the mouse model of NAFLD, treatment with CSP significantly reduced body weight gain and serum TG levels of the mice, and high-dose CSP treatment resulted in obvious reduction of ALT levels and hepatic fat accumulation. Network pharmacology analysis identified quercetin and 2-monolinolenin as the key bioactives in CSP, which target TNF, AKT1, IL6, TP53, and ALB. Docking simulations suggested strong binding between the two core compounds and their target proteins. The results of qRT-PCR showed that high-fat diet induced significant downregulation of Tp53, Cpt1, and Ppara expressions in mice, which was effectively reversed by CSP treatment. CONCLUSIONS CSP can improve lipid metabolism disorders in NAFLD mice through a regulatory mechanism involving multiple targets and pathways to reduce liver fat accumulation and protect liver function. The key components in CSP such as quercetin and linolenic acid monoacylglycerol may participate in the regulation of such metabolic processes as fatty acid oxidation by targeting TP53.
Animals ; Non-alcoholic Fatty Liver Disease/drug therapy* ; Mice ; Drugs, Chinese Herbal/pharmacology* ; Lipid Metabolism/drug effects* ; Molecular Docking Simulation ; Disease Models, Animal ; Liver/metabolism* ; Male ; Lipid Metabolism Disorders/drug therapy* ; PPAR alpha/metabolism* ; Mice, Inbred C57BL ; Network Pharmacology

OBJECTIVES: To investigate the regulatory role of the NLRP3 signaling pathway in hepatocyte pyroptosis in nonalcoholic steatohepatitis (NASH) under hypoxia. METHODS: Twenty-four male C57BL/6 mice were randomized equally into hypoxic control (A), hypoxic NASH model (B), hypoxic NASH+NLRP3 inhibitor (C), and hypoxic NASH+caspase-1 inhibitor (D) groups. In groups B-D, the mice were fed a methionine choline-deficient (MCD) diet under hypoxic conditions (to simulate a 5000 m altitude) for 6 weeks; the mice in groups C and D received intraperitoneal injections of the respective inhibitors every other day. RESULTS: Compared with those in group A, the mice in group B showed significantly elevated serum levels of FBG, TC, TG, ALT and AST, increased liver lipid content, inflammatory cell infiltration and collagen fiber deposition, and enhanced hepatic expressions of NLRP3, caspase-1, IL-1β and GSDMD proteins, with obvious swelling, cristae breakage, vacuolization, and outer membrane disruption of the mitochondria, ribosome loss in the cytoplasm, destruction of the nuclear membrane, and pathological changes of the rough endoplasmic reticulum. Treatment with NLRP3 inhibitor and caspase-1 inhibitor both significantly lowered serum levels of TC, TG, ALT and AST (but without significantly affecting FBG) in the mouse models, and reduced liver lipid content, inflammatory cell infiltration, collagen deposition, and expression levels of NLRP3, caspase-1, GSDMD and IL-1β. The treatments also significantly improved pathological changes in the mitochondria, ribosomes and endoplasmic reticulum in liver tissues of the mice. CONCLUSIONS NLRP3 signaling pathway plays a key role in promoting hepatocyte pyroptosis in NASH mice under hypoxic condition, and inhibiting this pathway can effectively reduce liver inflammation, suggesting its potential as a therapeutic target for NASH treatment.
Animals ; Non-alcoholic Fatty Liver Disease/metabolism* ; NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; Pyroptosis ; Mice, Inbred C57BL ; Male ; Hepatocytes/pathology* ; Signal Transduction ; Mice ; Hypoxia/metabolism* ; Caspase 1/metabolism* ; Interleukin-1beta/metabolism* ; Liver/metabolism*

OBJECTIVE: Prunella vulgaris L. has long been used for liver protection according to traditional Chinese medicine theory and has been proven by modern pharmacological research to have multiple potential liver-protective effects. However, its effects on non-alcoholic steatohepatitis (NASH) are currently uncertain. Our study explores the effects of P. vulgaris polysaccharides on NASH and intestinal homeostasis. METHODS: An aqueous extract of the dried fruit spikes of P. vulgaris was precipitated in an 85% ethanol solution (PVE85) to extract crude polysaccharides from the herb. A choline-deficient, L-amino acid-defined, high-fat diet (CDAHFD) was administrated to male C57BL/6 mice to establish a NASH animal model. After 4 weeks, the PVE85 group was orally administered PVE85 (200 mg/[kg·d]), while the control group and CDAHFD group were orally administered vehicle for 6 weeks. Quantitative real-time polymerase chain reaction analysis, Western blotting, immunohistochemistry and other methods were used to assess the impact of PVE85 on the liver in mice with NASH. 16S rRNA gene amplicon analysis was employed to evaluate the gut microbiota abundance and diversity in each group to examine alterations at various taxonomic levels. RESULTS: PVE85 significantly reversed the course of NASH in mice. mRNA levels of inflammatory mediators associated with NASH and protein expression of hepatic nucleotide-binding leucine-rich repeat and pyrin domain-containing protein 3 (NLRP3) were significantly reduced after PVE85 treatment. Moreover, PVE85 attenuated the thickening and cross-linking of collagen fibres and inhibited the expression of fibrosis-related mRNAs in the livers of NASH mice. Intriguingly, PVE85 restored changes in the gut microbiota and improved intestinal barrier dysfunction induced by NASH by increasing the abundance of Actinobacteria and reducing the abundance of Proteobacteria at the phylum level. PVE85 had significant activity in reducing the relative abundance of Clostridiaceae at the family levels. PVE85 markedly enhanced the abundance of some beneficial micro-organisms at various taxonomic levels as well. Additionally, the physicochemical environment of the intestine was effectively improved, involving an increase in the density of intestinal villi, normalization of the intestinal pH, and improvement of intestinal permeability. CONCLUSION PVE85 can reduce hepatic lipid overaccumulation, inflammation, and fibrosis in an animal model of CDAHFD-induced NASH and improve the intestinal microbial composition and intestinal structure. Please cite this article as: Zhu MJ, Song YJ, Rao PL, Gu WY, Xu Y, Xu HX. Therapeutic role of Prunella vulgaris L. polysaccharides in non-alcoholic steatohepatitis and gut dysbiosis. J Integr Med. 2025; 2025; 23(3): 297-308.
Animals ; Non-alcoholic Fatty Liver Disease/drug therapy* ; Male ; Dysbiosis/drug therapy* ; Mice, Inbred C57BL ; Gastrointestinal Microbiome/drug effects* ; Polysaccharides/therapeutic use* ; Prunella/chemistry* ; Mice ; Liver/metabolism* ; Plant Extracts/therapeutic use* ; Disease Models, Animal ; Diet, High-Fat

Obesity and metabolic dysfunction-associated steatotic liver disease (MASLD) are linked to numerous chronic conditions, including cardiovascular disease, atherosclerosis, chronic kidney disease, and type II diabetes. Previous research identified the natural flavonoid diosmin, derived from Chrysanthemum morifolium, as a regulator of glucose metabolism. However, its effects on lipid metabolism and underlying mechanisms remained unexplored. The AMP-activated protein kinase (AMPK) pathway serves a critical function in glucose and lipid metabolism. The relationship between diosmin and the AMPK pathway has not been previously documented. This investigation examined diosmin's capacity to reduce lipid content through AMPK pathway activation in hepatoblastoma cell line G2 (HepG2) and 3T3-L1 cells. The study revealed that diosmin inhibits lipogenesis, indicating its potential as an anti-obesity agent in obese mice. Moreover, diosmin demonstrated effective MASLD alleviation in vivo. These findings suggest that diosmin may represent a promising therapeutic candidate for treating obesity and MASLD.
Diosmin/administration & dosage* ; Animals ; AMP-Activated Protein Kinases/genetics* ; Humans ; Non-alcoholic Fatty Liver Disease/enzymology* ; Mice ; Obesity/enzymology* ; Hep G2 Cells ; Male ; 3T3-L1 Cells ; Mice, Inbred C57BL ; Signal Transduction/drug effects* ; Lipid Metabolism/drug effects* ; Chrysanthemum/chemistry* ; Lipogenesis/drug effects*